Fatty acid composition for treatment of alzheimer&#39;s disease and cognitive dysfunction

ABSTRACT

This invention relates to the use of a fatty acid composition comprising at least (all-Z omega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivatives thereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA), or derivatives thereof for manufacturing of a medicinal product or a food stuff for the treatment and/or prevention of amyloidos-related diseases, such as Alzheimer&#39;s disease, as well as treatment/prevention of cognitive dysfunction.

TECHNICAL FIELD OF THE INVENTION

The present invention relates to a fatty acid composition for the use inmedicinal products as well as in food stuff or food complements fortreatment and/or prevention of amyloidos-related diseases, such asAlzheimer's, and/or cognitive dysfunction. Additionally, the presentinvention relates to methods for treating and/or preventingamyloidos-related diseases, such as Alzheimer's, and/or cognitivedysfunction, which method comprises administering to a patient atherapeutically effective amount of a fatty acid composition.

BACKGROUND ART

The biological function of a protein depends on its three-dimensionalstructure, which is determined by its amino acid sequence during theprocess of protein folding. Normal folding is needed for successful cellfunctioning and therefore important in maintaining health. Several typesof diseases have been found where protein misfolding and conformationalchange are the main causes of appearance and progression of the diseases(1).

Misfolding of proteins may lead to formation of so called fibrils.Proteins or fragment of proteins are converted from their normallysoluble forms to insoluble fibrils or plaques, which accumulate in avariety of organs. The final forms of these aggregates often have awell-defined pathological anatomical appearance, known as amyloid.

Despite the range of proteins involved with their unique andcharacteristic native folds, the fibrils of the amyloid in which theyare found in the disease states are extremely similar in their overallappearance. Proteins known to have the propensity of fibrillarconformation in humans, called precursor proteins, are making up a listof 21 exponents (3) and the number is increasing. Usually the protein inthe fibril is made up of a small number of amino acids on average around20-60 grouping them in the category of polypeptides rather thanproteins.

Proteins are usually made up of an alfa-helix and a beta-sheet. Amyloidfibrils, however, usually contain beta-sheet material only rendering themolecules physical properties different from the parent protein. Whilenormal proteins are subjected to a continuous process of degradation byproteolysis, one very important feature of fibrils is the ability, onceformed, to be essentially indestructible under physiological conditions.The amyloid fibrils are dominated by hydrogen bonding between the amidand the carbonyl groups of the main chain, rather than by specificinteractions of the side chains, which determine the structure of normalproteins. This abnormal bonding induced by the large number of hydrogenbonds of the beta-sheet that must be disrupted to rescue the polypeptidechain from the aggregated state, results in a high resistance todegradation and properly removal from the tissue of deposition.

While in alfa-helices the hydrogen bonds are between side groups withinthe same strand, in beta sheets the bonds are between one strand andanother. Since the second beta-strand can come from a different regionof the same protein or from a different molecule, formation ofbeta-sheets is usually stabilised by protein oligomerisation oraggregation. In this manner the misfolded protein self-associates andbecome deposited in amyloid aggregates in diverse organs, inducingtissue damage and organ dysfunction. An important part of the depositionprocess is that a critical concentration of the precursor proteins hasto be present before fibril formation occurs (4). It also seems that assoon as an amyloid nucleus has been created the process of aggregationand deposition of amyloid material escalates.

Many of the precursor proteins are not directly prone to fibrildeformation. However, when peptide fragments of the precursor proteindissociate from the parent molecule such peptides do not have a stableglobular fold to protect them against aggregation. Folding of proteinsis a function of physical properties inherent from the amino acidsequence of the chain. These so called non-covalent interactions areweak bonding forces, however, the large number of individual contactswithin a protein adds up to a large energy factor favouring normalprotein folding. The most important force is the hydrophobic interactionbut even hydrogen bonds mentioned above are extremely important.Examples of even weaker forces are electrostatic interactions and vander Waals forces. The number of non-covalent interactions is to somedegree a function of the protein chain length meaning that splicing of asection of the protein to a peptide will render the peptide with lessstability due to the lower number of non-covalent interactions. Thenormal folding forces will be weaker which could favour the formation offibrils.

Less known but significantly important for normal folding as well asmaintenance of a stable three-dimensional structure, is proteinacylation by covalent attachment of fatty acids (5). It is wellestablished that the protein albumin is able to bind several moleculesof fatty acids. Saturated fatty acids such as stearic, palmitic andmyristic acid are the predominant fatty acids that attach to proteins ineukaryotic cells (6). From studies using radiolabelled fatty acids weknow that each fatty acid labels a different sub-population of proteinswith the fatty acid interacting with basic amino acids such as lysine,glycine and arginin. The carboxyl group of the fatty acid forms a saltbridge or a hydrogen bond with basic amino acid side chains. All siteshave cylindrical hydrophobic channels of varying shape that force thesaturated fatty acids to assume a nearly linear configuration. However,the binding pockets are large enough to accommodate unsaturated fattyacids such as oleic acid and arachidonic acid (7).

Interestingly, established amyloid also contains a certain amount offatty acids. By methanol extraction of amyloid derived fromtransthyretin about 10% of the dry mass was soluble pointing to thepresence of a lipid fraction (8). Gas-chromatography revealed thepresence of mixtures of saturated fatty acids like those mentionedabove, but also to polyunsaturated fatty acids like palmitoleic acid,linoleic acid, alfa-linolenic acid and arachidonic acid. This pattern offatty acids is typical for a modern Western diet, which is very muchbased on saturated fat from dairy products and meat together with seedderived oils. It is quite clear that fatty acids have a function in thenormal folding of proteins. The reason why fatty acids are found inamyloid is obscure but interestingly enouth the fatty acids found arecongruent with the fats of our diet. One hypothesis is based on theassumption that some fatty acids bound to the polypeptide or proteinhave weaker affinity rendering the chain less stable and therefore proneto fibrillar deformation.

Amyloid deposits can be reabsorbed and organ function reversed if thesynthesis of amyloidogenic protein is shut down. There seems to be afine balance between the rate at which amyloid is formed and itsclearance. It may therefore be possible to promote the resorption ofamyloid by reducing the concentration of the amyloidogenic protein to alevel below a critical threshold without necessarily eliminating theprecursor (AA). Studies of the mechanism of conversion from normallysoluble precursor proteins into amyloid fibrils have benefited from thefact that the transition can be reproduced under laboratory conditions.In vitro experiments have demonstrated that conversion of native, fullyfolded protein into a highly amyloidogenic, partially folded conformercould be blocked by stabilizing native proteins with a specific ligand(9). Other experiments using native precursor proteins such astau-protein (10) and islet amyloid polypeptide (IAPP) (11) have shown astimulating effect of certain fatty acids on the assembly of fibrils andamyloid. All long-chain fatty acids tested enhanced assembly to someextent, although greater stimulation was associated with unsaturatedforms. Both articles concluded that polyunsaturated fatty acids such asarachidonic acid, oleic acid and linoleic acid but also myristic acidexerted pronounced effects on fibril and amyloid formation. It seemedtherefore that common unsaturated fatty acids in our diet couldstimulate the formation of fibrils and amyloid and consequently increasethe risk of inducing disabling diseases like Alzheimer's disease.

SUMMARY OF THE INVENTION

Based on the present invention a number of aspects are presented in theappended claims. These aspects include, but are not limited to:

1. Use of a fatty acid composition for the treatment and/or preventionof amyloidos-related diseases, such as Alzheimer's disease.

2. Use of a fatty acid composition for the treatment and/or preventionof cognitive dysfunction.

3. Use of a food stuff or food supplement comprising a fatty acidcomposition for the treatment and/or prevention of amyloidos-relateddiseases, such as Alzheimer's disease.

4. Use of a food stuff or food supplement comprising a fatty acidcomposition for the treatment and/or prevention of cognitivedysfunction.

5. A method for treatment and/or prevention of amyloidos-relateddiseases, such as Alzheimer's disease.

6. A method for treatment and/or prevention of cognitive dysfunction.

As used herein, the term “Alzheimer's disease” is to be taken to includeboth Alzheimer's disease, Alzheimer's dementia and Alzheimer'ssclerosis.

A common feature for the above aspects is the use of a fatty acidcomposition comprising at least a combination of (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA), orderivatives thereof.

According to a first aspect of the invention, the invention relates tothe use of a fatty acid composition comprising at least a combination of(all-Z omega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), orderivatives thereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoicacid (EPA), or derivatives thereof for the production of a medicinalproduct for the treatment and/or prevention of amyloidos-relateddiseases, such as Alzheimer's disease.

This first aspect also relates to the use of said fatty acid compositionin treatment of amyloidos-related diseases, such as Alzheimer's disease,as well as a medicinal product for treatment of Alzheimers's diseasecomprising said fatty acid composition.

From research leading to the invention it was surprisingly found that afatty acid composition according to the invention prevents formation ofso called fibrils or plaques, and/or reduces deposed fibrils or plaques,known as amyloid. Moreover, a fatty acid composition according to theinvention predominantly containing DHA and EPA, or derivatives thereof,seemed to prevent and/or delay formation of fibrils most effectively,wherein the combination of DHA and EPA may act as an antagonist.

Moreover, the treatment according to the invention could be preventivereducing the propensity of fibril formation as well as therapeutic insituations with established amyloid.

Further, under unfavourable conditions, proteins or fragment of proteinsare converted from their normally soluble forms to insoluble fibrils orplaques, which accumulate in a variety of organs including the liver,kidneys, spleen, brain, and internal secretory glands like the betacells of the pancreas inducing toxic effects on cells and tissue. Thefinal forms of these aggregates often have a well-defined pathologicalanatomical appearance, known as amyloid. This is the reason for the useof the term amyloidoses to describe many of the clinical conditions withwhich deposition of amyloid are associated. Thus, as used herein, theterm “amyloidos-related” diseases means clinical conditions or diseaseswith which deposition of amyloid, preferably as a consequence of fibrilformation, are associated, such as for instance Alzheimer's disease.

In a second aspect, the present invention relates to the use of a fattyacid composition comprising at least a combination of (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA),or derivatives thereof, for the production of a medicinal product forthe treatment and/or prevention of cognitive dysfunction. This secondaspect also relates to the use of said fatty acid composition in thetreatment of cognitive dysfunction, as well as a medicinal product fortreatment of cognitive dysfunction comprising said fatty acidcomposition.

It was found that the fatty acid composition of the present inventionwas effective in decreasing the rate of memory decline in patientssuffering from cognitive dysfunction, for example in patients sufferingfrom mild cognitive dysfunction.

In a preferred embodiment of both the above first and second aspects ofthe present invention, the invention relates to the use of a fatty acidcomposition, wherein the weight ratio of EPA:DHA in the fatty acidcomposition is 1:2 to 2:1.55.

In another preferred embodiment of both the above first and secondaspects of the present invention, the invention relates to the use of afatty acid composition, wherein the weight ratio of EPA:DHA in the fattyacid composition is 1:X, where X is equal or greater than 1. Please notethat X being one of an integer or non-integer.

The term “amount” herein relates to weight or volume of the fatty acidcomposition.

In addition, the invention also includes a fatty acid compositioncomprising a combination of one or more DHA and/or EPA derivatives.Moreover, the desired pharmacological and/or therapeutic effect may beachieved by the fatty acid composition according to the invention.

In a preferred embodiment of the invention, EPA and DHA in the fattyacid composition are present in the composition in an EPA:DHA ratio from1:1 to 1:8. In a more preferred embodiment the EPA:DHA ratio in thefatty acid composition is from about 1:1 to 1:6. In a further embodimentof the invention, the fatty acid composition is a product containing DHAor a DHA-derivative in combination with EPA or an EPA-derivative.

Moreover, in another embodiment, the fatty acids in the compositionaccording to the invention is presented in at least one of esterifiedform, ethyl ester form, salt form, phospholipid form and free acid form,or any combinations thereof. In a preferred embodiment, the fatty acidcomposition is comprised of a combination of EPA and DHA in triglycerideform.

In another embodiment, at least DHA is obtained from at least one ofvegetable, microbial and animal origins, or combinations thereof.Moreover, in a further embodiment, wherein the fatty acid compositioncomprising at least a combination of DHA and EPA, at least one of DHAand EPA is obtained from at least one of vegetable, microbial and animalorigins or combinations thereof. The medicinal product or pharmaceuticalproduct includes therefore for instance a fatty acid compositioncomprising at least one of a DHA-containing microbial oil and a mixtureof an DHA-containing oil from microbial origin and a EPA-containing oilfrom a marine origin. Moreover, the fatty acid composition according tothe invention may additionally also comprise at least one of arachidonicacid (ARA), docosapentaenoic acid, heneicosapentaenoic acid andoctadecatentraenoic or derivatives thereof, or any combinations thereof.Suitably, at least a part of the EPA and/or DHA is produced from amarine oil, preferably a fish oil. Furthermore, in another embodiment ofthe medicinal product, the fatty acid composition is produced from amarine oil, such as a fish oil.

In a specific embodiment of the invention, the fatty acid composition iscomprised of at least a combination of EPA and DHA in triglyceride form.Moreover, it should be pointed out that the fatty acid composition isadministered to a human or an animal, preferably orally.

In a specific embodiment of the invention, the medicinal product is inform of a capsule or contained in a sachet. Preferably, the capsule isflavoured. More preferably, the capsule is a gelatine capsule which isflavoured. This embodiment also includes a capsule, therein both thecapsules and the encapsulated fatty acid composition are flavoured. Byflavouring the capsule as mentioned above, the capsule becomes moreattractive to the user. However, a medicinal product according to theinvention may also be produced for administration though any other routewhere the active ingredients may be efficiently absorbed and utilized,e.g. intravenously, subcutaneously, intramuscularly, intranasally,rectally, vaginally or topically.

In another embodiment, said fatty acid composition is administered in anamount providing a daily dosage of 1 g to 15 g of said fatty acidcomposition for a human. In a preferred embodiment between 2 and 10 g ofsaid fatty acid composition is administered per day, and in a morepreferred embodiment between 2 and 8 g of said fatty acid composition.The medicinal product or pharmaceutical composition or pharmaceuticalpreparation according to the invention may also comprise othersubstances such as an inert vehicle, or pharmaceutical acceptableadjuvants, carriers, preservatives etc., which are well known in tothose skilled in the art. However, the medicinal product may also beadministered to an animal, such as a pet or a horse. Moreover, it shouldbe pointed out that the medicinal product may be at least one of anamyloid-preventing agent or amyloid-deposit decreasing agent.

In another embodiment of the invention, the invention relates to a useof a fatty acid composition comprising at least (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA), orderivatives thereof, that upon administration to a human or an animalwill prevent the formation of aggregates of protein fibrils or plaqueand/or reduce deposed fibrils, for the production of a medicinal productfor the treatment and/or prevention of amyloidos-related diseases, suchas Alzheimer's disease, and/or cognitive dysfunction.

According to a third aspect of the invention, the present inventionrelates to the use of a fatty acid composition comprising at least(all-Z omega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), orderivatives thereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoicacid (EPA), or derivatives thereof, for the production of a food stuffor food supplement for the treatment and/or prevention ofamyloidos-related diseases, such as Alzheimer's disease.

From research leading to the invention it was surprisingly found that afatty acid composition according to the invention prevents formation ofso called fibrils or plaques, and/or reduces deposed fibrils or plaques,known as amyloid. A fatty acid composition containing at least acombination of the two fatty acids DHA and EPA seemed to prevent and/ordelay formation of fibrils most effectively. Moreover, the use accordingto above leads to the same advantages and possibilities as mentionedbefore. The treatment according to the invention could be preventive,reducing the propensity of fibril formation, as well as therapeutic insituations with established amyloid. The definition concerning“amyloidos-related” diseases is also included for the patent positionsrelated to a food stuff or a food supplement according to the invention.

In a fourth aspect, the present invention also provides the use of afatty acid composition comprising at least (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA),or derivatives thereof, for the production of a food stuff or foodsupplement for the treatment and/or prevention of cognitive dysfunction.

It was found that the fatty acid composition of the present inventionwas effective in decreasing the rate of memory decline in patientssuffering from cognitive dysfunction, for example in patients sufferingfrom mild cognitive dysfunction.

One advantage of manufacturing and selling a food stuff for thetreatment and/or prevention of amyloidos-related diseases, such asAlzheimer's disease, or for treatment and/or prevention of cognitivedysfunction is that such a food stuff will be more easily accessible forpeople. In preventive purpose they preferably buy the product orsupplement in a health store and/or a supermarket, and they do not needto visit a doctor.

In a preferred embodiment of the third and fourth aspects of the presentinvention, the invention relates to the use of a fatty acid composition,wherein the weight ratio of EPA:DHA in the fatty acid composition is 1:2to 2:1.55.

In another preferred embodiment of the third and fourth embodiments ofthe present invention, the invention relates to the use of a fatty acidcomposition, wherein the weight ratio of EPA:DHA in the fatty acidcomposition is 1:X, where X is equal or greater than 1. Please note thatX being one of an integer or non-integer. In addition, the inventionalso includes a fatty acid composition comprising at least a combinationof one or more DHA and/or EPA derivatives. Moreover, the desiredpharmacological and/or therapeutic effect may be achieved by the fattyacid composition according to the invention.

In another embodiment of the invention, EPA and DHA in the fatty acidcomposition are present in the composition of an EPA:DHA ratio from 1:1to 1:8. In a more preferred embodiment the EPA:DHA ratio in the fattyacid composition is from about 1:1 to 1:6. In a specific embodiment ofthe invention, the fatty acid composition is a product of DHA or aDHA-derivative and EPA or an EPA-derivative.

Moreover, in another embodiment, the fatty acids in the compositionaccording to the invention is presented in at least one of esterifiedform, ethyl ester form, salt form, phospholipid form and free acid form,or any combinations thereof. In a preferred embodiment, the fatty acidcomposition is comprised of a combination of EPA and DHA in triglycerideform.

Further, in another embodiment, at least DHA is obtained from at leastone of vegetable, microbial and animal origins, or combinations thereof.In preferred embodiment, wherein the fatty acid composition comprisingat least a combination of DHA and EPA, at least one of DHA and EPA isobtained from at least one of vegetable, microbial and animal origins orcombinations thereof. The food stuff or food supplement includestherefore, for instance, a fatty acid composition comprising at leastone of a DHA-containing microbial oil and a mixture of an DHA-containingoil from microbial origin and a EPA-containing oil from a marine origin.Further, the fatty acid composition according to the invention mayadditionally also comprise at least one of arachidonic acid (ARA),docosapentaenoic acid, heneicosapentaenoic acid and octadecatentraenoicor derivatives thereof, or any combinations thereof. Suitably, at leasta part of the EPA and/or DHA is produced from a marine oil, preferably afish oil. Furthermore, in another embodiment of the food stuff or foodsupplement, the fatty acid composition is produced from a marine oil,such as a fish oil.

In a preferred embodiment of the invention, the fatty acid compositionis comprised of at least a combination of EPA and DHA in triglycerideform. Moreover, it should be pointed out that the fatty acid compositionis administered to a human or an animal, preferably orally. However, thefood stuff or food supplement according to the invention may also beproduced for administration though any other route as mentioned before.

In a specific embodiment of the invention, the food stuff or foodsupplement is in form of a capsule or contained in a sachet. Preferably,the capsule is flavoured. More preferably, the capsule is a gelatinecapsule which is flavoured. This embodiment also includes a capsule,therein both the capsules and the encapsulated fatty acid compositionare flavoured. By flavouring the capsule as mentioned above, the capsulebecomes more attractive to the user.

In another preferred embodiment, said fatty acid composition isadministered in an amount providing a daily dosage of 1 g to 15 g ofsaid fatty acid composition for a human. In a more preferred embodimentbetween 2 and 10 g of said fatty acid composition is administered perday, and in a most preferred embodiment between 2 and 8 g of said fattyacid composition. The food stuff or food supplement according to theinvention may also comprise other substances such as an inert vehicle,or pharmaceutical acceptable adjuvants, carriers, preservatives etc.,which are well known in to those skilled in the art. Additionally, eventhe food stuff or food supplement may be administered to an animal suchas a pet or a horse. Moreover, it should be pointed out that the foodstuff or food supplement may be at least one of an amyloid-preventingagent or amyloid-deposit decreasing agent.

In a specific embodiment, the present invention relates to use of afatty acid composition comprising at least DHA, or derivatives thereof,and EPA, or derivatives therof, that upon administration to a human oran animal will prevent the formation of aggregates of protein fibrils orplaque and/or reduce deposed fibrils, for the production of a medicinalproduct for the treatment and/or prevention of amyloidos-relateddiseases, such as Alzheimer's disease, and/or cognitive dysfunction.Moreover, one advantage of selling a food stuff for prevention ofAlzheimer's disease or a food stuff for preventing cognitive dysfunctionis that the food stuff may help people not to develop these diseases inthe future.

According to a fifth aspect of the invention, the present inventionrelates to a method for the treatment and/or prevention ofamyloidos-related diseases, such as Alzheimer's disease, wherein aneffective amount of a fatty acid composition comprising at least acombination of (all-Z omega-3)-4,7,10,13,16,19-docosahexaenoic acid(DHA), or derivatives thereof, and (all-Zomega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA), or derivativesthereof, is administered to a human or an animal. Herein, “an effectiveamount” also includes a therapeutically or a pharmaceutically activeamount of the fatty acid composition. This expression relates to a doseof said fatty acid composition that will lead to the desiredpharmacological and/or therapeutic effect. The desired pharmacologicaland/or therapeutic effect is, as stated above, achieved by the fattyacid composition according to the invention. From research leading tothe invention it was surprisingly found that a fatty acid compositionaccording to the invention prevents formation of so called fibrils orplaques, and/or reduces deposed fibrils or plaques, known as amyloid.

In a sixth aspect, the present invention also relates to a method forthe treatment and/or prevention of cognitive dysfunction, wherein aneffective amount of a fatty acid composition comprising at least acombination of (all-Z omega-3)-4,7,10,13,16,19-docosahexaenoic acid(DHA), or derivatives thereof, and (all-Zomega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA), or derivativesthereof, is administered to a human or an animal.

It was found that the fatty acid composition of the present inventionwas effective in decreasing the rate of memory decline in patientssuffering from cognitive dysfunction, for example in patients sufferingfrom mild cognitive dysfunction.

In a preferred embodiment of the fifth and sixth aspects of the presentinvention, the invention relates to the use of a fatty acid composition,wherein the weight ratio of EPA:DHA in the fatty acid composition is 1:2to 2:1.55. In another preferred embodiment of the fifth and sixth aspectof the invention, the fatty acid composition the weight ratio of EPA:DHAin the fatty acid composition is 1:X, where X is equal or greaterthan 1. Moreover, the method leads to the same advantages andpossibilities as mentioned before.

In a preferred method for treatment of Alzheimer's disease and/orcognitive dysfunction according to the invention, the disease is causedby deposition of amyloid. In a specific embodiment of the invention, themethod of treating Alzheimer's disease ans/or cognitive dysfunction isrelated to administering to a human suffering from the disease aneffective disease (Alzheimer's and/or cognitive dyfunction) alleviatingamount of a amyloid-deposit-decreasing agent. Preferably, theamyloid-deposit-decreasing agent is a fatty acid composition comprisingat least one of a combination of the fatty acids DHA, derivativesthereof, and EPA, derivatives thereof.

In another embodiment of the invention, the embodiment relates to use ofa fatty acid composition comprising at least a combination of DHA, orderivatives thereof, and EPA, or derivatives thereof, that uponadministration to a human or an animal will lead to prevention of, orreduction of, deposition of amyloid, for the manufacture of a medicamentor a food supplement, for the treatment and/or prevention of Alzheimer'sdisease, preferably caused by amyloidosis.

In a specific embodiment of the invention, the fatty acid composition ofthe present invention, for use in a medical product, food stuff, foodsupplement or method of treatment comprises at least 70% unsaturatedomega-3 fatty acids wherein the fatty acids DHA and EPA are present in aweight ratio from about 1:2 to 2:1.

As used herein, amyloidos-related conditions or diseases, include forexample Alzheimer's disease. Such diseases/conditions can be sporadic,inherited or even infectious, and often occur only late in life even ifinherited forms may appear much earlier. Each disease is associated witha particular protein and aggregates of these proteins are thought to bethe direct origin of the pathological conditions associated with thedisease. Moreover, the treatment and/or prevention according toadministering the fatty acid composition of the invention may alsoinclude at least one of; treatment due to reduction of amyloidaggregates, prevention of misfolding of proteins that may lead toformation of so called fibrils or plaque, treatment due to decreasing ofthe production of Aβ-protein (amyloid beta protein), and preventionand/or treatment due to inhibiting or slow down the formation of proteinfibrils, aggregates, or plaque. Moreover, the present invention alsoincludes prevention of fibril accumulation, or formation, byadministering a fatty acid composition according to the invention.

In addition, as used herein the term “treatment” means both treatmenthaving a curving or alleviating purpose and the treatment of aamyloidos-related disease, such as Alzheimer's, or cognitive dysfunctioncan be made either acutely or chronically. By chronically treatment ismeant a treatment witch continues for weeks or years.

BRIEF DESCRIPTION OF THE DRAWINGS

In the studies and examples below reference is made to the accompanyingdrawings. Herein, reference is made to the accompanying drawings onwhich:

FIG. 1 shows fibril formation of omega-3 preparations during 30 minutes;

FIG. 2 demonstrates fibril formation between the fatty acid preparationswith the most prominent fibril inducing effect, soy oil and olive oil,compared to the fibril preventing DHA-concentrate EPAX 2050 (whichcomprises approximately 20% EPA and 50% DHA);

FIG. 3 shows preventive effect against fibril formation up to at least150 minutes comparing different omega-3 preparations;

FIG. 4 illustrates effects against fibril formation with aDHA-concentrate compared to olive oil and soy oil; and

FIG. 5 illustrates results from one of the experiments below, showingthe effect on the MMSE-scoring by orally administering EPAX® 2050TG topatients suffering from cognitive dysfunction.

DESCRIPTION OF PREFERRED EMBODIMENTS

A number of preferred embodiments of the invention were performed inorder to demonstrate that a fatty acid composition comprising acombination fo DHA and EPA is effective on the treatment and/orprevention of amyloidos-related diseases.

Fibril formation is a consequence of misfolding the precursor proteins.The reason for this abnormal behaviour of forming normalthree-dimensional structures has not been fully elucidated. As discussedin the background art, it could seem that common polyunsaturated fattyacids frequently recommended by dieticians to prevent cardio-vasculardisease and cancer, could in fact increase the propensity of misfoldingprecursor proteins, thereby inducing amyloid deposition. However, thepresent invention disclose results showing that another marinelong-chain polyunsaturated fatty acid, namely docosahexaenoic acid(DHA), surprisingly seemed to indicating a preventive effect on amyloidformation by prolonging the time to spontaneous fibril formation ofIAPP.

The present invention discloses the results of experiments with onesynthetic precursor protein, synthetic IAPP, spontaneously formingfibrils. This peptide was incubated with free fatty acids of marineorigin comparing the effects on fibril formation with fatty acids likeoleic acid and linoleic acid known to stimulate the formation of amyloidfibrils. Moreover, the present invention also discloses results ofexperiments in a semi-vivo model, where pancreatic islets fromtransgenic mice producing human IAPP and spontaneously forming beta-callamyloid were prepared and incubated with oleic acid and docosahexaenoicacid.

In a first preferred embodiment, the effects of different omega-3preparations on fibril formation, were studied.

In a second embodiment, the effects of a fatty acid composition rich inDHA, EPAX 2050® (a high omega-3 oil), an oil comprising at least acombination of DHA and EPA, (K85: approximately 460 mg EPA and 375 mgDHA), and an olive oil, on deposition of amyloid fibrils in pancreaticislets, were studied.

EXAMPLES

In Vitro Experiments on Fibril Formation

In the first study, the effects on fibril formation of different omega-3preparations were studied. The in vitro fibril formation experimentswere performed in small glass test tubes. A stock of dissolved fattyacids in ethanol was prepared by dissolving fatty acids in 100% ethanolat a concentration of 10 mM. After mixing with the same amount ofconcentrated NaOH, the final concentration of ethanol was 3%. The fattyacids or fatty acid combinations in the table below were tested. 10 mMFatty acids Mol. weight contained Linoleic acid 278 27.8 mg (fromsoybean oil) Oleic acid 280 28.0 mg (from olive oil) EPAX DHAconcentrate, EPAX2050 301 30.1 mg (≈500 mg DHA/g and ≈200 mg EPA/g) EPAX4510 318 31.8 mg (≈450 mg EPA/g) EPA 95 302 30.2 mg (≈950 mg EPA/g) K85314 31.4 mg (≈465 mg EPA/g and ≈375 mg DHA/g) Control synthIAPP +ethanol

Synthetic precursor protein, synthetic IAPP spontaneously formingfilbrils, was dissolved in dimethylsulfoxide (DMSO) at a concentrationof 10 mg/ml. 25 μM was incubated with each one of the fatty acids, eachone in a concentration of 125 μM, in distilled water. One μl aliquots ofeach sample were analysed after 5, 10, 20, 30, 60, 90, 120, 150, 180,210, and 240 minutes in the electron microscope after negativecontrasting with 2% uranyl acetate in 50% ethanol. Formation of fibrilswas observed by electron microscopical analysis and recorded as scoresarbitrarily between 0 and 5 in the 30 minutes experiments and between 1and 2 in the 240 minutes experiments. FIG. 1 demonstrates fibrilformation of the omega-3 preparations during 30 minutes. The omega-3concentrate containing predominantly DHA, EPAX 2050, seemed to preventor delay the spontaneously formation of fibrils most effectively.Moreover, FIG. 2 demonstrates fibril formation between the fatty acidpreparation with the most prominent fibril inducing effect, olive oiland soy oil, compared to the fibril preventing DHA-concentrate EPAX2050.

In addition, fibril induction was also followed for 240 minutes toquantify the fibril preventive effect of the DHA-concentrate. FIG. 3demonstrates that preventive effect against fibril formation was evidentup till 150 minutes in the experiment comparing different omega-3preparations. Similar prevention was obtained in the experiment with theDHA concentrate compared to olive oil and soy oil, see FIG. 4.

Thus, this study shows that treatment with a fatty acid composition richin DHA leads to prevention of fibril or plaque formation. Moreover, itseams that DHA act as an fibril inhibitor. At the same time theinvention also shows preventive effect on fibril formation of a productcomprising at least a combination of DHA and EPA, wherein preferably theamount of DHA≧EPA. The results also suggest a specific preventive effectagainst fibril formation of an omega-3 product of marine origincomprising EPA and DHA as compared with soy oil and olive oil.

In Vitro Experiments on Amyloid Formation in Pancreatic Islets

In the second study protective effects on amyloid deposition of an oliveoil, a high DHA omega-3 oil, and a composition comprising at least DHAand EPA, wherein the amount of EPA≧DHA, were studied.

Transgenic mice carrying the human IAPP gene may be used for studyingdeposition of amyloid fibrils in the pancreatic islets (11). Therefore,single pancreatic islets were isolated and cultured from transgenicmice. Pancreas were removed under sterile conditions and placed inHank's balanced salts and finely minced. Small pieces of tissue wereenzymatically digested by collagenase for 10 minutes in a 37 degreeCelcius water bath. The islets were individually selected under themicroscope. Subsequently the islets were cultured overnight in 24 wellcell cluster containing RPMI 1640 medium supplemented with 10% fetalbovine serum, penicillin (100 U/ml), streptomycin (1, 1 mg/ml) and 22, 0mM glucose at 37 degrees in humidified air containing 5% carbonmonoxide.

500 μM of olive oil, high DHA omega-3 oil (EPAX 2050) and K85 oil and 1%fatty acid-free albumin were added to the wells and the islets werecultured in RPMI medium during 2 weeks. There were about 60 islets ineach well. Congo red from a stock solution was added to the wells andthe islets were examined using light microscopy. Amyloid deposition isstained with Congo red while other cell material does not.

No difference was observed between the different islet groups regardingsurvival (Table 1). No. of living islets Fatty acid Start End EPAX 205060 50 K 85 60 52 Olive oil 65 54

The results of this study shows that 4-5% of the cell islets incubatedin the olive oil solution were stained with Congo red indicatingintracellular amyloid deposition. Surprisingly only 1% of the cells fromislets incubated with high DHA, but even also the K85 concentrate, werestained with Congo red indicating a protective effect against amyloiddeposition. However, the DHA rich product, EPAX 2050TG, exhibitedstronger effect compared to K85 (EPA≧DHA). Additionally, the results ofthis study confirm effects supporting prevention and treatment ofamyloidos-related diseases influenced of a fatty acid compositioncomprising at least DHA and EPA, or derivatives thereof.

Prevention of Cognitive Decline

The aim of this study was to evaluate the effects of high dose omega-3fatty acid supplementation to patients suffering from Alzheimer'sdisease (AD-patients) in preventing cognitive decline and Alzheimer'sdisease.

From year 2000 to 2004, 204 AD-patients with a mean age 74±9 (range54-90) years, under choline esterase inhibitor treatment, were blockrandomised to either 4 grams daily of omega-3 fatty acids intriglyceride form (˜2.8 g DHA and ˜1.2 g EPA, EPAX® 2050TG, PronovaBiocare) or placebo (4 g linoleic acid) for 6 months.

After 6 months, all participants received active treatment for 6 moremonths. Only patients with Mini Mental State Examination (MMSE) >15points and with no other serious illness were included.

Of the 204 patients participating in the study, 174 patients (73±9 y,52% women) fulfilled the protocol and are reported here. The drop outrate (n=30) did not differ between the two treatment arms.

Mean MMSE initially in all patients were 23.5±4 points for the group toinitially receive treatment and 27±10 points in the group to initiallyreceive placebo.

In the whole group, the decline in cognitive function, i.e MMSE˜−0.8±2.6points, did not differ according to randomisation group at the 6-monthfollow-up.

However, in the subgroup of patients with mild or very mild cognitivedysfunction, i.e. MMSE >27 points, there was a significant reduction inMMSE decline rate in the treatment group (Δ=−0.5±1.5 points, i.e. from28.4±0.9 to 27.9±1.9 points, n=19) over the first 6 month treatmentperiod in comparison with the placebo group (Δ=−2.6±2.8 points, i.e.from 28.6±1.1 to 26±2.1 points, n=12) (p<0.05 between groups).

A similar arrest in decline rate was observed in the placebo group(MMSE >27 p) when they received active omega-3 fatty acid treatmentbetween 6 and 12 months (from 26±2.1 to 25.2±3.1 p). The results for thepatient subgroup having initial MMSE >27 is shown in FIG. 5.

These results indicates that high doses of a combination of the omega-3fatty acids DHA and EPA, given to patients with mild or very mildcognitive dysfunction and/or mild or very mild Alzheimer's disease,delayed the rate of cognitive decline according to MMSE.

Discussion

In the studies presented above, the synthetic precursor protein IAPPspontaneously forming fibrils were incubated in vitro with a series offatty acids of the omega-3 series as well as omega-6 and omega-9. Theomega-3 concentrate containing DHA and EPA seemed to prevent or delaythe spontaneous formation of fibrils while the omega-6 (soybean oil) andomega-9 (olive oil) fatty acids seemed to provoke fibril formation. Thelast pattern is known from previous experiments (10, 11). The preventiveeffect of the combination of DHA and EPA, however, is an unexpectedfinding. Other omega-3 fatty acids did not have the same effect asDHA:EPA even if some preventive effects were observed compared to theomega-6 and 9 oils. Moreover, in the amyloid in vitro model on isletsfrom transgenic IAPP producing mice DHA, but even another highomega-concentrate, K85, induced amyloid deposition to a significant lessextent compared to the omega-9 olive oil.

The present findings indicate a novel therapeutic modality to diseasescaused by amyloid deposition. The treatment could be preventive reducingthe propensity of fibril formation as well as therapeutic in situationswith established amyloid.

Finally, the results support the use of a medicinal product, apharmaceutical composition, a food stuff or a food supplement,comprising a fatty acid composition comprising a combination of DHA andEPA, for the treatment and/or prevention of amyloidos-related diseases,such as for instance Alzheimer's disease, as well as for cognitivedysfunction.

The invention shall not be limited to the shown embodiments andexamples.

REFERENCES

-   1) Zerovnik A. Eur J Biochem 2002; 269:3362-3371-   2) Merlini G and Belotti V. NEJM 2003; 349:583-596-   3) Westermark P, Benson M D, Buxbaum J N, et al. Amyloid 2002;    9:197-200-   4) McLaurin J, Yang D, Yip C M, et al. J Struct Biol 2000;    130:259-270-   5) Schmidt M F G. Biochim. Biophys. Acta 1989; 988:411-426-   6) Mclhinney R A J, Cdadwixk J K, and Pelly S J. Biochim. 1987;    244:109-115-   7) Bhattacharya A A. J Mol Biol 2000; 303:721-732-   8) Hermansen L F, Bergman T, Jörnvall H, et al. Eur J Biochim 1995;    227:772-779-   9) Miroy G J, Lai Z, Lashuel H A, et al. Proc Natl Acad Sci USA    1996; 93:15051-15056-   10) Wilson D M and Binder L I. Am J Pathol 1997; 150:2181-2195)-   11) Ma Z and Westermark G T. Medical Dissertation No. 655,    Linkoping, Sweden 2001

1.-33. (canceled)
 34. A method for treatment or prevention of anamyloidos-related disease, wherein said disease is Alzheimer's diseaseby administering a therapeutically effective amount of a fatty acidcomposition including at least (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA),or derivatives thereof, for the production of a medicinal product, foodstuff or food supplement.
 35. A method for treatment and/or preventionof cognitive dysfunction by administering a therapeutically effectiveamount of a fatty acid composition comprising at least (all-Zomega-3)-4,7,10,13,16,19-docosahexaenoic acid (DHA), or derivativesthereof, and (all-Z omega-3)-5,8,11,14,17-eicosapentaenoic acid (EPA),or derivatives thereof, for the production of a medicinal product, foodstuff or food supplement.
 36. The method of treatment or preventionaccording to claim 34, wherein the combination of EPA and DHA arepresent in the composition in an EPA:DHA ratio from about 1:2 to 2:1.55.37. The method of treatment or prevention according to claim 34, whereinthe weight ratio of EPA:DHA in the fatty acid composition is 1:X, whereX is equal or greater than
 1. 38. The method of treatment or preventionaccording to claim 37, wherein the combination of EPA and DHA arepresent in the composition in an EPA:DHA ratio from 1:1 to 1:8,preferably in an EPA:DHA ratio from 1:1 to 1:6.
 39. The method oftreatment or prevention according to claim 34, wherein the fatty acidsin the composition is presented in at least one of esterified form,ethyl ester form, salt form, free acid form and phospholipid form or anycombinations thereof.
 40. The method of treatment or preventionaccording to claim 34, wherein at least DHA is obtained from at leastone of vegetable, microbial and animal origins or combinations thereof.41. The method of treatment or prevention according to claim 34, whereinat least one of DHA and EPA is obtained from at least one of vegetable,microbial and animal origins or combinations thereof.
 42. The method oftreatment or prevention according to claim 34, wherein at least a partof the EPA and/or DHA is produced from a marine oil, preferably a fishoil.
 43. The method of treatment or prevention according to claim 34,wherein the fatty acid composition is produced from a marine oil. 44.The method of treatment or prevention according to claim 34, wherein thefatty acid composition is comprised of at least a combination of EPA andDHA in triglyceride form.
 45. The method of treatment or preventionaccording to claim 34, wherein the fatty acid composition isadministered orally to a human or an animal.
 46. The method treatment orprevention according to claim 34, wherein said fatty acid composition isadministered in an amount providing a daily dosage of 1 g to 15 g ofsaid fatty acid composition, preferably between 2 and 6 g for a human.47. The method of treatment or prevention according to claim 34, whereinsaid medicinal product, food stuff or food supplement is at least one ofan amyloid-preventing agent and/or an amyloid-decreasing agent.
 48. Themethod of treatment or prevention according to claim 35, wherein saidcognitive dysfunction is a mild cognitive dysfunction.
 49. The method oftreatment or prevention according to claim 34, wherein theadministration to a human will prevent the formation of aggregates ofprotein fibrils or plaque and/or reduce deposed fibrils.
 50. The methodof treatment or prevention according to claim 34, wherein said fattyacid composition is contained in a oral dosage form selected from thegroup consisting of a capsule and a sachet.
 51. The method of treatmentor prevention according to claim 50, wherein said oral dosage form isflavoured.
 52. The method of treatment or prevention according to claim45, wherein the administration to a human will prevent the formation ofaggregates of protein fibrils or plaque and/or reducedeposed fibrils.